Vertical movement of microplastics by roots of wheat plant (Triticum aestivum) and the plant response in sandy soil

Abstract

Microplastics persist as a challenging pollutant in agroecosystems, posing potential risks to soil health and crop productivity. Root growth, elongation and expansion may significantly influence the vertical transport and infiltration of microplastics into the soil profile. Wheat plants (Triticum aestivum) grown in 70 cm deep rhizotrons were investigated for their influence on the vertical movement of two prevalent microplastic shapes, polyester fibres and polyvinyl chloride (PVC) fragments. Wheat was chosen for its dense and extensive fibrous and fine root system, which is a robust model for studying root-soil-microplastic interactions. Microplastics at a 0.24% w/w dry soil weight concentration were homogeneously distributed in the topsoil (0–20 cm). Infiltration of polyester fibres up to 50 cm into the soil profile was discerned as strong adherence to plant roots. PVC fragments exhibited greater mobility, reaching depths of 70 cm in the presence and absence of wheat plants. Plant growth response on exposure to microplastics appeared in the form of increased root branching and decreased shoot biomass, indicating a stress response in wheat plants. The results prove the vertical movement of microplastics, while the infiltration depth was influenced by microplastic shape. Movement was detected as either strong adherence of polyester fibres to plant roots or infiltration of PVC fragments. PVC fragments may have infiltrated through preferential flow paths in soil pores and the fissures created by root elongation and water movement.

Introduction

Microplastics are generated from meso- and macro-plastics breakdown and are less than 5 mm in size [1]. With microplastic particles accumulating in the soils, there is growing concern over their potential risk to agroecosystems [2]–[4]. In agroecosystems, microplastic particles occur primarily as fibres [5 – 8], and fragments [9]; [10]; [7]. Their main sources are irrigation with treated wastewater [11], the introduction of compost and the use of plastic mulching [9]; [12]; [13]. Zhang et al. [10] determined that approximately 92% of all microplastics in their agricultural soil saples were fibres, while the remaining 8% primarily consisted of fragments and films.

The vertical and horizontal movement of microplastics through the soil profile has been investigated for the influence of soil cracking, water infiltration, runoff, tillage, digging mammals, and earthworms [14–16]. Taylor et al. [17] observed microplastic movement because of root growth and found that 40 nm and 1 μm polystyrene spheres accumulated on plant root tips grown in gel cultures. In aquatic plants, Mateos-Cárdenas et al. [18] and Kalčíková et al. [19] determined that 10–45 μm polyethylene microplastics and 30–600 μm polyethylene microbeads respectively, adhered to the entire root surface, consequently moving during root elongation and expansion. Despite this, the influence of root growth and elongation on microplastic transport was not the primary focus of these studies. Li et al. [20] is the only study to investigate the role of plant roots on microplastic movement, concluding that the downward movement of 1–2 mm polypropylene films and beads was due to water infiltration, while their upward movement was due to plant root growth. Moreover, the positioning of microplastics in the soil profile determined their influence [20]. Initial deposition on the soil surface leads to minimal vertical migration by crop roots, while uniform distribution along the topsoil profile (up to 30 cm) increases their tendency to be transported to its middle layers [20]. The correlation between microplastic particle shape and their transport by roots remains largely unexplored.

In porous media and due to water flow, particle shape would determine the degree of entanglement in the pores [21], where spherical and irregularly shaped particles have been shown to experience greater mobility than fibres and fragments [22– 24]. Additionally, the intensity of entanglement in porous media would be higher with fibres than fragments [22– 24] [21]. Zubris et al. [8] emphasised the potential persistence of fibres detected in long-term soil studies 15 years after the last application of organic sewage sludge from wastewater, with evidence of vertical movement along preferential flow paths. Hu et al. [25] found microplastic fibres and fragments at a depth of 80 cm and 30 cm, respectively, after 10 years of continuous plastic mulching, while after 32 years of continuous plastic mulching, Li et al. [26] found fibres at a depth of 100 cm. Regarding land use, Qiu et al. [11] found microplastic fibres at a depth of 100 cm and fragments at a depth of 50 cm in frequently saturated paddy fields where rice with fibrous roots is grown. In contrast, fibres were found to infiltrate to 80 cm and fragments to 20 cm in land irrigated by treated wastewater and experienced wetting and drying periods [11]. The microplastic shape plays a pivotal role in the movement, whereas their polymer type and density effect seem negligible [23].

This research study hypothesises that the shape and type of microplastic particles and plant root growth and expansion significantly influence the vertical infiltration depth and distribution of microplastics in soil. Laboratory experiments were conducted where wheat (Triticum aestivum) was grown in rhizotrons under the same hydrological conditions. The wheat plant was chosen for its fast life cycle, compact size, and suitability for growth in a climate chamber. In contrast, its dense fibrous root system is ideal for studying root-soil-microplastic interactions. Each rhizotron was exposed to two different microplastic shapes (fibres and fragments). Polyester fibres and polyvinyl chloride (PVC) fragments, with documented fragmentation-dependent vertical transport due to water [27], were utilised in this current study. Polyester fibres and PVC fragments were selected due to their prevalence and differing physical characteristics, as well as their chemical stability and environmental persistence for long-term studies. PVC fragments were selected over polyester fragments to explore the impact of different polymer types on infiltration behaviour under similar hydrological conditions. Furthermore, the consequent plant response to exposure to microplastics was evaluated. This study aims to offer initial indications of the microplastic size- and shape-dependent transport in the root zone of soil.

Materials and methods

Sand and microplastics preparation

The quartz sand in the planting medium was prepared according to Tumwet et al. [27]. in the 1.0–2.0 mm size range. The quartz sand was dry, clean and free of organic compounds and fine fractions. The soil was presumed to have high permeability, whereas its lack of organic compounds would reduce the degree of soil aggregation and biofilm formation. Red polyvinyl chloride (PVC) microplastic fragments were prepared and characterised according to Tumwet et al. [27]. The resulting particles were characterised as 125–300 μm hydrophobic fragments (density; 1.35 g / cm³, “Technoplast”, purchased from Treskow GmbH^®, Germany) (Fig. S1). Red polyester microplastic fibres were generated by manually cutting 100% polyeter wool “Happy Chunky Double” (product number 17,880, Hobbii A/S Copenhagen, Denmark, density; 1.2 g / cm³) with sterilised scissors. The fibres had an average length of 5000 ± 300 μm and a diameter of 21 ± 2 μm. They were characterised as hydrophobic fibres (Fig. S2). In this study, microplastics resemble those found in the environment where secondary microplastics are produced by mechanical abrasion (i.e., grinding of a PVC pipe).

2.4 g of microplastics and 1000 g of sand were mixed in a glass beaker by stirring the mixture with a sterilised metal spoon for 15 min to create a uniform mixture. The microplastic-sand mixture level of 0.24% $\:w/w$ dry soil weight simulated here was within the range (0.1 × 10⁵ – 2.0% $\:w/w$ dry soil weight) of contaminated soils near anthropogenically affected landscapes [28]. The microplastic-soil mixture was homogenously mixed in the top 20 cm of the rhizotron (MS_mix) soil profile, replicating microplastic exposure as it would likely occur in farmland soils [5]. Therefore, approximately 1.15 × 10⁶ polyester fibres and 3.39 × 10⁹ PVC fragments were initially added.

Experimental design

The experiment was conducted in a climate chamber at the University of Applied Sciences, Zittau/Görlitz (HSZG). The temperature in the climate chamber was set to 22 °C (during the day, 14 h) and 17 °C (at night, 10 h) with a constant relative humidity of 70%. A full spectrum artificial light (35.2 µmol/s, Bioledex^® GoLeaf LED Plant-tube full spectrum) was used as the light source above each rhizotron. The rhizotron opening is 42 cm long and 5 cm wide, and its 76 cm by 42 cm walls are held on by bolts that can be easily opened at the end of the experimental period. 24.32 kg of sand would, therefore, fill a rhizotron. Rhizotrons were then filled up to 51 cm with sand, and the top 20 cm was filled with the MS_mix. 15 rhizotrons were set up in the climate chamber.

Leibniz Institute of Plant Genetics and Crop Plant Research supplied wheat seeds (Triticum aestivum) via the Standard Material Transfer Agreement. Wheat seeds (0.04–0.05 g) of an almost uniform size were selected; their surface was not sterilised. 9 seeds were sowed directly into each rhizotron at a depth of 5 cm and approximately 4.5 cm apart. After 7 days, thinning was conducted and 6 uniformly sized seedlings were retained in the rhizotrons. In 3 rhizotrons (RZ1_{control, wheat}, RZ2_{control, wheat} and RZ3_{control, wheat}), 6 plants were planted in each. The sand in these rhizotrons did not contain microplastics. Accordingly, they were used as the control group to investigate how the wheat plants would grow without microplastics. To examine how wheat roots would alter the infiltration behaviour of microplastics, MS_mix with polyester was added in 6 rhizotrons, where 3 (RZ4_{polyester, wheat}, RZ5_{polyester, wheat} and RZ6_{polyester, wheat}) had 6 plants grown in each, while in a control 3 (RZ7_{polyester, control}, RZ8_{polyester, control} and RZ9_{polyester, control}), no wheat was grown in them. Additionally, MS_mix with PVC fragments were added in 6 rhizotrons in which 3 (RZ10_{PVC, wheat}, RZ11_{PVC, wheat} and RZ12_{PVC, wheat}) had 6 plants in each, and 3 (RZ13_{PVC, control}, RZ14_{PVC, control} and RZ15_{PVC, control}) had no plants and used as a control. Thus, 18 plants were exposed to polyester fibres, 18 were exposed to PVC fragmenting particles, and 18 were grown without microplastics. A total of 54 wheat plants were investigated for the role of plant roots in the vertical movement of microplastics. Table 1 shows the configuration of each rhizotron. The rhizotron used and a schematic representation of the experiment setup (Fig. S3) are provided in the supplementary material. The rhizotrons were watered with hydroponics nutritive solution (pH: 5.8, General hydroponics Maxi Gro™): 100 ml daily from Monday to Thursday and 200 ml on Fridays. The experiment was terminated just before the flowering stage of wheat plants at 118 days to explore the microplastic infiltration depth. All 54 plants survived.

Table 1 Table of the configuration of each rhizotron. (RZ = rhizotron)

Microplastic extraction and quantification

At the end of the experimental period, the rhizotrons were placed on their side, and the 76 cm by 42 cm wall was opened. Sand sampling was done at specific depths of 2.5 cm, 5 cm, 10 cm, 20 cm, 35 cm, 50 cm, and 65 cm. Across each depth, 6 sand samples were scooped out by a sterilised metal spoon, carefully excavating around the roots to avoid disturbing the attached microplastics to collect 240 g of the sand and microplastic mixture. Polyester fibres were then classified by their characteristics, such as colour, size, and shape, in ten steps, accounting for 24 g of the sand and microplastics mixture at a time. PVC fragments were extracted according to Tumwet et al. [27]. This involved density separation with CaCl₂ solution (prepared from Calcium chloride dihydrate p.a., min. 99.5%, Chemsolute^®, density ≈ 1.4 g/cm3) coarse sand and vacuum filtration with a 20 μm nylon net filter paper (Merck Millipore^®). Visual identification and quantification of the red polyester fibres and PVC fragments were carried out using microscopy, and their mass was recorded on a scale (Sartorius analytical balance Practum^® 224–1 S, readability 0.1 mg, max. 220 g). As 3 rhizotrons (RZ1_{control, wheat}, RZ2 _{control, wheat} and RZ3 _{control, wheat}) served as blanks, the distinct colour, size, and shape of the investigated microplastics could easily be differentiated from atmospherically deposited and background microplastics.

The wheat plants were carefully lifted from the rhizotron still on its side, and excess sand was carefully shaken off. They were then placed on a piece of cardboard and allowed to air dry. Microplastics attached to the roots at the specific investigated depths were quantified by carefully using sterilised tweezers to separate the adhered microplastics from the roots. The microplastics were visually identified based on shape, colour, and size. The microplastics were weighed on a scale (Sartorius analytical balance Practum^® 224–1 S, readability 0.1 mg, max. 220 g).

Plant parameters measurements

Plant parameters such as the shoot and root length and biomass, the biomass of the leaves and their surface area were measured. The roots were photographed, and their surface area was determined using the ink method, according to Sattelmacher et al. [29]. In the ink method, roots were stained for 1 min with a 1:100 diluted solution of black drawing ink (Pelikan©, 5-chloro-2-methyl-2 H-isothiazol-3-one and 2-methyl-2 H-isothiazol-3-one (3:1)) and distilled water. Excess ink was removed by carefully rotating the plant and left to drain for 5 s, and adhering ink was allowed to diffuse into distilled water (50 or 100 ml according to root size). The quantity of ink was determined photometrically (photometer SPECORD^®) at 500 nm in a cuvette with 2 ml of sample. A spectrometry was carried out, and the optical density was measured. Optical density is linearly correlated with root surface area, allowing for a relative comparison of the surface areas. Two calibration lines were created for the evaluation (Fig. S4).

QA/QC

A comprehensive Quality Assurance and Quality Control (QA/QC) procedure was implemented throughout the study to ensure the reliability and validity of the experimental results. Blank rhizotrons (RZ1_{control, wheat}, RZ2 _{control, wheat} and RZ3 _{control, wheat}), without microplastics, were included to check for potential microplastic contamination from external sources during sample preparation, handling, and analysis. Positive control samples containing known quantities of polyester fibres and PVC fragments were prepared by adding the specified amounts of fibres and fragments to the soil and then analysed using the same methods as the experimental samples to validate the extraction, detection and measurement methods. Each experimental condition had three replicates to assess variability and ensure consistency in the results. This included six separate rhizotrons for each microplastic shape (fibres and fragments). All analytical instruments, including balances, microscopes, and density separation solution, were calibrated regularly according to manufacturer guidelines to ensure accurate measurements. Standardised methods were used for sampling, handling, and analysing microplastics. This included consistent procedures for introducing microplastics into the soil, watering the plants, and measuring microplastic adherence. Detailed records of all experimental procedures, observations, and any deviations from the planned methods were maintained. This provided a clear audit trail and allowed for thorough review and verification of the results. Environmental variables such as temperature, humidity, and light conditions were carefully controlled and monitored throughout the experiment to minimise their impact on the results. Appropriate statistical methods, including one-way and two-way ANOVA, were used to analyse the data. This ensures that the results are statistically significant.

Statistical analysis

All the data underwent an initial screening for outliers through scatter plots and experimental records. The data was then adjusted to ensure a normal distribution and uniform variances. Two-way ANOVA was conducted to analyse how the shape of microplastic particles and soil depth interact to influence their vertical infiltration. Summary tables of the two-way ANOVA results considering the depth and plant, i.e., Plant 1, Plant 2, Plant 3, etc. on the quantity of microplastics in rhizotrons are provided in Table S1 and Table S2. One-way ANOVA was conducted to examine the effects of microplastic type on root biometrics, leaf size, shoot size, and their respective biomass. Additional ANOVA tests were performed to compare wheat plants exposed to PVC and polyester against the control group. Python 3.10^® was used for statistical analyses. The data’s normality was assessed using the Shapiro–Wilk test for normal distribution, and a significance level (α) of 0.05 was used. A Post Hoc (t-test) was conducted when statistically significant differences were identified. A table showing this statistical evaluation of plant parameters is included in the supplementary material (Table S3).

Results and discussion

Influence of root growth on vertical transport of microplastics

The wheat plant roots were studied at the end of the experimental period to quantify the microplastics adhered to them and their occurrence depth. Sand samples collected at 2.5 cm, 5 cm, 10 cm, 20 cm, 35 cm, 50 cm, and 65 cm were investigated for the microplastic infiltration depth. The red polyester fibres and PVC fragments were identified and quantified. Figure 1 shows the appearance of reanimated roots in the control group (plants grown in the absence of microplastics, RZ1-3_{control, wheat}) (a) and those exposed to polyester fibres (RZ4-6_{polyester, wheat},) (b) and PVC fragments (RZ10-12_{PVC, wheat}) (c).

Fig. 1

Elongated fibrous root system of the wheat roots. Roots in the control (a), in the presence of (b) polyester fibres, and (c) PVC fragments. Secondary root systems were evident in roots not exposed to microplastics. These three roots were dried and reanimated in a water cylinder 50 cm long to display the apparent different root systems

Upon seed germination, a primary root system developed with a secondary root system observed in nodes above the primary one in plants in the control while sparse in those in the soil substrate with microplastic, i.e., MS_mix. The root system was noticeably finer and more extensive with exposure to microplastics, with roots exposed to PVC particles exhibiting even increased branching degrees.

In the 3 blank rhizotrons (RZ1_{control, wheat}, RZ2_{control, wheat} and RZ3_{control, wheat}), 6 plants were planted in each without microplastics. At the end of the experimental period, an average of 5 microplastic fibres of various lengths, primarily blue, were found in each rhizotron. The contamination was possibly from external sources during sample preparation, handling, and analysis. No fragments were found in the rhizotrons. The number distribution across the depth is represented in Table S4.

Polyester fibres

Polyester-sand mix was added and homogenised in 6 rhizotrons at a concentration of 2.4 g microplastics per 1000 g of sand soil in the first 20 cm (MS_mix). The infiltration depth of the polyester was investigated in the presence and absence of fibrous wheat root plants. At the end of the experimental period, the wheat plants in RZ4_{polyester, wheat}, RZ5_{polyester, wheat} and RZ6_{polyester, wheat} were carefully removed from the planting medium, air dried, and the polyester adhering to them quantified. The mass of polyester occurring at different depths of the sand was also quantified. A summary of the polyester (mass in g) at different sampled infiltration depths in sand soil in the presence of wheat plants is presented in Fig. 2.

Fig. 2

Polyester infiltrating in the presence of wheat in RZ4_{polyester, wheat}, RZ5_{polyester, wheat} and RZ6_{polyester, wheat}. The average mass of polyester quantified in each rhizotron is presented. Error bars represent standard error. The sampling depth (cm) is not to scale but a representation of the specific sampling points

With MS_mix comprising the sand up to a depth of 20 cm, polyester was found in the sand at a depth of 2.5 cm, 5 cm, 10 cm, and 20 cm in RZ4_{polyester, wheat} and RZ5_{polyester, wheat}, while RZ6_{polyester, wheat} had less polyester in the sand and none occurring in the samples at 2.5 cm and 10 cm. Some of the polyester quantified in the sand soil in RZ4_{polyester, wheat} noticeably had dead roots firmly attached to them that the tweezers couldn’t separate. No polyester was found deeper than 20 cm after careful wheat root removal after the 118-day experimental period. This suggests limited vertical movement of polyester fibres beyond the middle soil layers, likely due to their shape and interaction with the soil and root structure. The highest concentration of polyester fibres was found at 10 cm (0.679 g) and 20 cm (0.762 g) in RZ4_{polyester, wheat}, with notable amounts at 2.5 cm (0.54 g) and 5 cm (0.42 g). This suggests that polyester fibres are more concentrated in the upper and middle soil layers. Polyester fibres were also present in significant quantities at 2.5 cm (0.516 g) and 5 cm (0.304 g) in RZ5_{polyester, wheat}, but showed lower overall concentrations than in RZ4_{polyester, wheat}. At 10 cm, the concentration was 0.354 g; at 20 cm, it was 0.432 g. Fibres were absent at 2.5 cm but present at 5 cm (0.294 g) and 20 cm (0.391 g) in RZ6_{polyester, wheat}, indicating some vertical movement but less accumulation compared to RZ4_{polyester, wheat} and RZ5_{polyester, wheat}.

The polyester adhering to the roots of each of the 6 plants grown in RZ4_{polyester, wheat}, RZ5_{polyester, wheat} and RZ6_{polyester, wheat} was quantified and presented in Fig. 3. The roots of Plant 1, Plant 2, Plant 3, Plant 4, Plant 5, and Plant 6 are hereafter referred to as PR1, PR2, PR3, PR4, PR5, and PR6 respectively. A summary of the quantity of polyester occurring attached to the plant roots at each investigated depth is provided in the supplementary material (Table S5).

Fig. 3

Quantity of polyester adhering to the roots of each of the 6 plants in RZ4_{polyester, wheat}, RZ5_{polyester, wheat} and RZ6_{polyester, wheat}. The polyester quantity on each plant root system is represented by its corresponding plant colour. After removing the wheat plants, polyester in the soil was observed up to 20 cm, where RZ4_{polyester, wheat} had the highest sampled amount. Error bars represent standard error. The sampling depth (cm) is not to scale but a representation of the specific sampling points

RZ4_{polyester, wheat} had the highest quantity of polyester occurring in the sand, and it was found that more polyester remained in the soil than adhered to the roots. During sampling, some dead roots were found firmly attached to the polyester fibres within the top 20 cm of the soil profile. PR2, PR3 and PR6 had no polyester adhered to them throughout their entire length. Up to 20 cm, the MS_mix depth, the average ratio of the quantity of polyester adhering to the roots (PR1, PR2 and PR5) to those in the soil was 1:10. Only PR1 and PR5 had fibres adhered to them at 35 cm weighing 0.091 g and 0.032 g respectively. PR1 had the highest quantity of polyester at each investigated depth. In RZ5_{polyester, wheat}, all 6 plants had polyester adhering to their roots at various depths. At 2.5 cm, 5 cm, 10 cm, 20 cm, 35 cm, and 50 cm, the average polyester adhering to the roots was 0.02 ± 0.008 g. In RZ6_{polyester, wheat}, each plant had polyester adhering to its roots at various depths, with polyester occurring in PR3, PR5 and PR6, deeper than their initial position (0–20 cm). Polyester was only found at 5 cm and 20 cm in the sampled soil.

The statistical analysis of polyester fibre adherence to wheat roots provides crucial insights into the relationship between microplastic shape, vertical infiltration depth, and plant growth. The significant effect of depth (p-value = 0.000016) demonstrates that fibre adherence varies notably across diverse soil depths, indicating that factors such as soil density, moisture, and root density at varying depths play pivotal roles. The significant differences among individual plants (p-value = 0.005883) highlight that specific plant characteristics, including root morphology and physiological traits, impact microplastic adherence. However, the non-significant interaction between depth and plant (p-value = 0.594276) suggests that the pattern of fibre adherence change with depth remains consistent across different plants. Therefore, the density of roots in upper soil layers provides more surface area for microplastic adherence than in deeper layers, while variations in root morphology and soil depth significantly impact the adherence of polyester fibres to plants.

In RZ7_{polyester, control}, RZ8_{polyester, control} and RZ9_{polyester, control}, homogenously mixed polyester and sand were added in the top 20 cm of the soil profile, and the infiltration behaviour of the polyester in the absence of the wheat plant roots was scrutinised. A summary of the mass of polyester occurring at various depths is presented in Fig. 4.

Fig. 4

Quantity of polyester infiltrating in sand in the absence of wheat roots in RZ7_{polyester, control}, RZ8_{polyester, control} and RZ9_{polyester, control}. The average mass of polyester quantified in each rhizotron is presented. Error bars represent standard error. The sampling depth (cm) is not to scale but a representation of the specific sampling points

Compared to RZ4_{polyester, wheat}, RZ5_{polyester, wheat} and RZ6_{polyester, wheat}, the average ratio of polyester quantity occurring in RZ7_{polyester, control}, RZ8_{polyester, control} and RZ9_{polyester, control} was 1:2, i.e. twice as much polyester was found in the samples in the absence of wheat roots. Polyester did not infiltrate past the initial position in which they were placed (0–20 cm).

The infiltration scenario of polyester in the sand was expected to be altered in the presence of roots. The polyester fibres firmly adhered to the fibrous wheat roots and infiltrated up to 35 cm in RZ4_{polyester, wheat} and 50 cm in RZ5_{polyester, wheat} and RZ6_{polyester, wheat}. In this experiment, the wheat plants were slowly and carefully lifted and removed at the end of the experimental period, extracting the polyester occurring at these depths. With the polyester firmly adhering to the roots, no polyester was found in the sand after their initial MS_mix position (0–20 cm) following the plants being uprooted. It was detected that polyester intertwined with the roots during the initial growth period, possibly intensifying with time. Harvesting in agricultural practices does not remove all wheat roots in the soil; thus, fibres would prevail to greater depths [26]. In each rhizotron, the planting position of the seedlings could affect the quantity of sampled polyester. Additionally, 3 seedlings were removed from the soil during thinning after 7 days. These seedlings showed apparent entanglement of the fibres, and the average seedling root length was 20 cm. More polyester was quantified in the soil in RZ4_{polyester, wheat} than in RZ5_{polyester, wheat}, with the least quantified in RZ6_{polyester, wheat}. With less polyester in the soil, it was observed that the polyester was more evenly distributed on the roots of the 6 plants in RZ6_{polyester, wheat} than in RZ4_{polyester, wheat} and RZ5_{polyester, wheat}. It was also observed that RZ6_{polyester, wheat} had the highest amount of polyester removed by the seedling roots at the initial 7 days, with RZ4_{polyester, wheat} having the least.

In each rhizotron with polyester fibres and in the presence of fibrous wheat root plants, polyester infiltrated past its initial position. The planting position might play a role in the entanglement of the microplastic fibres to the roots where, as early as 7 days, the seedling roots snared the polyester. This affected the quantity of polyester found after the plants were uprooted (118 days), as discerned in RZ6_{polyester, wheat}. Li et al. [26]. showed entanglement of fibrous roots and microplastics is prevalent and, depending on their size (0.2–2.0 μm), uptake of the microplastics by the wheat roots is possible [30].With water infiltration being the only possible factor influencing microplastic movement in RZ7_{polyester, control}, RZ8_{polyester, control,} and RZ9_{polyester, control}, entanglement in the soil pores could be a cause to impede movement [22]; [23]; [24]; [21]. Microplastic fibres have been found to have less mobility in porous media as they become intensely entangled in the pores [21].

In the wheat-grown rhizotrons (RZ4_{polyester, wheat}, RZ5_{polyester, wheat} and RZ6_{polyester, wheat},), polyester fibres were more concentrated in the upper and middle soil layers, especially noticeable in RZ4_{polyester, wheat} with the highest concentration at 20 cm (0.762 g). In control rhizotrons without wheat (RZ7_{polyester, control}, RZ8_{polyester, control} and RZ9_{polyester, control}), polyester concentrations were present but showed slightly different distribution patterns. RZ8_{polyester, control} had the highest concentrations across all depths, suggesting that polyester fibres settle more uniformly or are affected by soil compaction and gravity without root interference. Plant roots likely influenced the movement and retention of polyester fibres, with roots creating physical barriers that trap polyester fibres in wheat-grown rhizotrons, resulting in higher concentrations at specific depths. In control rhizotrons, polyester fibres may be more influenced by soil compaction and gravity, resulting in a relatively uniform distribution but potentially higher accumulation in certain spots due to soil settling.

The distribution of polyester fibres on the roots of wheat plants and in the surrounding soil at various depths provides valuable insights into the relationship between microplastics, plant roots, and soil profiles. The highest quantity of polyester fibres was found in the top layer (0–20 cm), indicating the initial even distribution of microplastics in the upper soil. This layer showed a substantial presence of fibres both adhered to roots and within the soil matrix. The strong attachment of fibres to plant roots in the upper layers suggests high surface area and root density, which facilitates capturing and retaining fibres, especially in plants with dense and fibrous root systems. The fibre concentration decreases with increasing depth, with considerable quantities still present in the 20–50 cm range. This distribution implies that fibres move vertically due to root growth and soil displacement. The adherence of fibres to roots at these depths highlights the role of root elongation and expansion in transporting fibres downward. Polyester fibres were less frequently found in the deeper soil layers (50–70 cm), indicating limited vertical movement beyond the mid-depths. The scarce presence of fibres in deeper layers also highlights the natural filtration ability of the soil and the diminishing effectiveness of root-mediated transport mechanisms at greater depths.

PVC fragments

2.4 g of PVC fragments were added to 1000 g of sand soil, uniformly mixed and placed in the first 20 cm of 6 rhizotrons (MS_mix). The infiltration depth of the PVC was investigated with (RZ10_{PVC, wheat}, RZ11_{PVC, wheat} and RZ12_{PVC, wheat}) and without (RZ13_{PVC, control}, RZ14_{PVC, control} and RZ15_{PVC, control}) fibrous wheat root plants. After the experimental period, the wheat plants in RZ10_{PVC, wheat}, RZ11_{PVC, wheat} and RZ12_{PVC, wheat} were carefully uprooted and air-dried, and the PVC attached to their roots were quantified. Additionally, the mass of PVC occurring at different depths of the sand was assessed. A summary of the PVC fragments (in grams) at different infiltration depths in sand soil in the presence of wheat plants is depicted in Fig. 5.

Fig. 5

PVC infiltration in the presence of wheat in RZ10_{PVC, wheat}, RZ11_{PVC, wheat} and RZ12_{PVC, wheat}. The average mass of PVC quantified in each rhizotron is presented. The PVC-sand mix was initially in the top 20 cm (0–20 cm). Error bars represent standard error. The sampling depth (cm) is not to scale but a representation of the specific sampling points

The MS_mix was up to a depth of 20 cm; as such, more PVC was quantified up to this depth. The highest levels of PVC were detected at 10 cm (0.486 g) and 5 cm (0.463 g) in RZ10_{PVC, wheat}, while in RZ11_{PVC, wheat}, the most significant concentration was found at 20 cm (0.377 g). In RZ12_{PVC, wheat}, the largest amount was found at 2.5 cm (0.619 g). The quantity of PVC infiltrating past 20 cm was quantifiably similar in all three rhizotrons, i.e., 0.01 g. This indicates limited vertical movement of PVC fragments beyond the middle soil layers, similar to the observations with polyester fibres. The PVC attached to the roots of each of the 6 plants grown in RZ10_{PVC, wheat}, RZ11_{PVC, wheat} and RZ12_{PVC, wheat} was quantified and shown in Fig. 6, with a summary in the supplementary material (Table S6).

Fig. 6

Quantity of PVC adhering to the roots of each of the 6 plants in RZ10_{PVC, wheat}, RZ11_{PVC, wheat} and RZ12_{PVC, wheat}. The quantity of PVC on each plant root system is represented by its corresponding plant colour. PVC in soil was observed the entire depth of the rhizotron. Error bars represent standard error. The sampling depth (cm) is not to scale but a representation of the specific sampling points

In RZ10_{PVC, wheat}, PVC concentrations were highest at 10 cm. The highest concentration of PVC was observed in PR1 at a depth of 10 cm, measuring 0.0024 g. In RZ11_{PVC, wheat}, the highest quantity of PVC fragments was found at a depth of 10 cm on PR1 (0.0068 g) and at a depth of 20 cm on PR4 (0.0096 g). Within RZ12_{PVC, wheat}, the PVC concentrations were generally lower compared to RZ11_{PVC, wheat}, but higher than RZ10_{PVC, wheat}, in some cases, with PR3 exhibiting the highest concentration (0.0036 g) at a depth of 2.5 cm. Minimal PVC concentrations were identified beyond a depth of 20 cm, implying limited vertical migration of PVC fragments beyond the upper layers, where root density and activity are more pronounced. Less PVC fragments were attached to the wheat roots than polyester fibres. PVC fragments were found in the entire depth of the rhizotron. Roots exposed to PVC exhibited increased degrees of branching.

The results indicate that both soil depth and individual plants have a significant impact on PVC attachment, with p-values of 0.005874 and 0.000244, respectively. This suggests that PVC fragment attachment varies significantly based on soil depth and the plant, emphasising the influence of vertical soil layers and plant root characteristics in interactions with microplastics. However, the lack of a significant interaction between depth and plant (p = 0.173432) indicates a consistent pattern in how soil depth affects PVC attachment across the different plants.

In the absence of wheat roots, the infiltration depth of PVC fragments was investigated. A summary of the mass of PVC occurring at various depths of the soil profile is presented in Fig. 7.

Fig. 7

Quantity of PVC infiltration in sand in the absence of wheat roots in RZ13_{PVC, control}, RZ14_{PVC, control} and RZ15_{PVC, control}. The average mass of PVC quantified in each rhizotron is presented. PVC fragments were found throughout the entire depth of the rhizotrons. Error bars represent standard error. The sampling depth (cm) is not to scale but a representation of the specific sampling points

The highest PVC concentration was found at 10 cm depth in RZ13_{PVC, control} (0.398 g) and at 2.5 cm depth in RZ14_{PVC, control} (0.415 g). RZ15_{PVC, control} had the highest concentration at 5 cm (0.305 g). Minimal PVC concentrations were detected beyond 20 cm, indicating limited vertical migration of PVC fragments in wheat-grown rhizotrons. Less PVC was found in the rhizotrons without wheat roots; however, this largely depends on the sampling position. Approximately 240 g of sand was collected at 6 positions across each investigated depth. After the initial MS_mix depth, the quantity of PVC infiltrating deeper was, on average, 0.01 g in the absence and presence of the roots at 35 cm, 50 cm and 65 cm. In rhizotrons without wheat, PVC concentrations were more evenly distributed within the 0–20 cm range. This suggests that the absence of plant roots allows for a more uniform settling of PVC particles, possibly due to soil compaction and gravity. Additionally, iwithout wheat roots, water infiltration could be the only possible factor influencing microplastic movement. Wheat-grown rhizotrons showed more significant variations in PVC concentrations at various depths, possibly because root growth created pathways or obstacles for PVC fragments. As the PVC was hydrophobic, adherence to roots would be less compared to hydrophilic microplastics, influencing how PVC fragments are immobilized within the root zone.

O’Connor et al. [24] determined that microplastics added on the surface (1.5 cm) of sand soil would be transported by water infiltration up to depths of 7.5 cm, and the microplastic size and pore space govern vertical movement as evidenced by Tumwet et al. [27] and Waldschläger et al. [23]. The infiltration patterns experienced by microplastics would be unimpeded infiltration, finite depth infiltration or surface sealing from clogged soil pores depending on the diameter of microplastic particles and the diameter of the porous media [27]; [23]. PVC was found in the entire depth of the rhizotron, indicating that the fragments infiltrated up to 70 cm within 118 days. The quantity of PVC infiltrating past 35 cm in the absence and presence of wheat plant roots was quantifiably similar.

Gao et al. [31] found that wet-dry cycles and a downward flow prompt vertical migration of microplastics influenced by gravity, and the higher the hydrophobicity, the less the infiltration [31]. Li et al. [20] established that microplastics (1–2 mm) in loam soil barely moved due to water infiltration, whereas the presence of crop roots prompted vertical migration. PVC used in this experiment was characterised as hydrophobic 125–300 μm fragments and wet-dry cycles were experienced during watering periods. Hydrophobic PVC fragments are less likely to adhere to water molecules, which reduces their likelihood of being carried along with water during infiltration. Therefore, an uneven distribution of PVC fragments is expected as preferential flow paths (i.e. cracks, macropores) are not uniform. During dry periods the formation of soil cracks is more likely, acting as conduits for rapid infiltration during subsequent wet periods. Also, PVC fragments are less likely to be drawn upward by capillary action during dry periods, leading to a more stable position within the soil profile than hydrophilic microplastics. As PVC fragments were found at significant depths, movement was through preferential flow paths rather than uniform infiltration. Furthermore, O’Connor et al. [24] found that the amount of microplastics quantified deeper in the soil profile after smaller infiltration events (50–100 ml) was typically more significant than those quantified after higher infiltration events (200–1000 ml). In this study, the rhizotrons were watered with 100 ml hydroponics solution daily from Monday to Thursday and 200 ml on Fridays. This likely allowed for slow percolation and vertical movement of the PVC through the soil profile.

Roots exposed to PVC exhibited noticeably increased branching degrees, possibly due to microplastics clogging the sand pore spaces and diverting the growth direction of the roots. Bao et al. [32] showed that overcoming root penetration resistance increases branching. It was deduced that the pores filled with water, therefore prompting secondary macropore flow paths in the root zone [33], and the presence of roots increased the pores in the soil that favoured the water infiltration path, therefore increasing the preferential flow paths [34]. Preferential flow paths were visible in these rhizotrons. PVC fragments possibly infiltrated through the fissures and biopores created by root elongation and growth.

It can be inferred that the movement of hydrophobic microplastics like polyethylene and polypropylene in the soil is expected to be confined to the upper layers due to their limited adhesion to water molecules. Conversely, hydrophilic microplastics are likely to infiltrate more uniformly owing to their stronger adhesion to water molecules and soil particles. Smaller and irregularly shaped microplastic fragments would have greater mobility than larger, uniform particles, as observed with PVC. Furthermore, plants with extensive, fibrous root systems (e.g., grasses and cereals) play a crucial role in establishing pathways for microplastic movement, as demonstrated with wheat roots.

Plant response to exposure to microplastics

Leaves

Leaf widths were, on average, similar for plants exposed to PVC and polyester microplastics, with leaves in the control group quantifiably broader. On average, leaves in the control group were longer than those exposed to microplastics. The average leaf surface area of plants exposed to PVC was the smallest at 75.4 cm², with those exposed to polyester being the largest at 87.4 cm^2, and the control surface area was, on average, 77 cm². On average, plants exposed to PVC and polyester microplastics and those in the control group had 7 leaves. A summary of the leaf width, length, surface area and leaf number per plant of wheat plants exposed to PVC and polyester microplastics and those in the control are presented in the supplementary material (Fig. S5). The results suggest a leaf width and length reduction due to microplastic exposure. Microplastics could impede root growth, alter the soil structure and aeration, disrupting nutrient and water availability and consequently induce stress responses in plants. However, the leaf surface area was highest in the plants exposed to polyester microplastic fibres. The reduced leaf surface area upon exposure to PVC fragments corroborates results from an experiment by Qi et al. [35], where a reduction in the leaf surface area of wheat (Triticum aestivum) was demonstrated when exposed to biodegradable microplastic particles (polyethylene terephthalate and polybutylene terephthalate) at 1.0% $\:w/w$ ry soil weight. Furthermore, Rillig et al. [36] discerned that microplastic fragments lead to a decreased leaf surface area compared to microplastic fibres.

The leaves of plants exposed to PVC had the lowest dry mass, while leaves in the control group had the highest (Fig. S6). The leaf dry mass in plants exposed to PVC fragments decreased by up to 14% compared to the control group, while the decrease in plants exposed to polyester fibres was 9%. The water content was, however, lowest in plants exposed to polyester, followed by the control and highest in plants exposed to PVC. Plants adapt to their root expansion obstructions, such as PVC fragmenting particles, by increasing water and nutrient uptake [37]. This circumstance might explain the higher water content in the leaves on exposure to PVC fragments. Exposure to polyester or PVC microplastics results in notable changes in leaf morphology and dry mass where PVC has a more pronounced negative impact, likely due to impaired root growth, altered soil structure, and disrupted nutrient and water availability. Microplastics, therefore, impact leaf health, with economic and ecological consequences for agriculture and plant communities. The allocated leaf biomass is depicted in Fig. 8.

Fig. 8

Total biomass (dry mass in grams) of the wheat plant roots, stem and leaves exposed to polyester and PVC (0.24% ω/ω soil) and a control. Error bars represent standard error. A post-hoc test was only performed when growth parameters were significantly affected by the occurrence of microplastics

Shoots

The shoot is the above-ground section of the wheat plant and includes the stem and leaves. The mean stem length was highest for plants in the control group, while the mean length for plants exposed to polyester was higher than that of plants exposed to PVC microplastics. On average, the stems of the plants exposed to PVC and polyester had, quantifiably, the same dry mass (Fig. 8). A summary of the stem length, dry mass, shoot dry mass, and water content is provided in the supplementary material (Fig. S7). Despite having the highest water content, shoots exposed to PVC were the shortest and had the lowest biomass. The results indicate a 20% reduced shoot biomass of plants exposed to microplastics and a 12% and 15% decrease in stem height of wheat plants exposed to polyester fibres and PVC particles, respectively. Future studies could explore other plant metrics, longer-term impacts, and different microplastic types. Examining varied soil conditions and environmental factors may provide more insights. The significant difference in variances should be considered for future studies to investigate underlying factors affecting plant growth variability that were not examined in this study.

These results are similar to those observed by Qi et al. [35]. Boots et al. [38], also observed a reduction in shoot height with biodegradable polylactic acid particles. On the contrary, Huang et al. [39] presumed that microplastic fragments in the soil positively affect the shoot biomass by reducing the abundance of harmful soil biota and promoting carbon mineralization and microbial activity. Shoots of plants exposed to PVC fragments had the highest water content, most likely due to increasing water uptake, an occurrence deduced by Mašková et al. [37]. However, de Souza Machado et al. [40], observed an increased water content of plants exposed to polyester fibres as fibres in the soil would significantly enhance water holding capacity of the soil. It was also concluded that microplastic size weakens wheat shoots, where microplastics are expected to exhibit a more negative impact than macroplastics, as Qi et al. [35] demonstrated.

It is evident that microplastic exposure reduces stem length, decreases shoot biomass, and alters water dynamics. This indicates stress induced by microplastics, which may arise from physical blockages, altered soil structure, or chemical interactions between the microplastics and soil matrix.

Roots

Polyester fibres firmly adhered to the root surface. The root surface area (Fig. S8) was calculated according to the ink method. Roots in the planting medium with polyester fibres had the highest surface area, 119% higher than those in the control. This is due to polyester adhering and ingrowing into the roots, forcing a more fibrous growth of the roots, which was observed. With polyester clinging to the roots, the roots grew the entire length of the rhizotrons. With this, the polyester infiltrated deeper than its initial occurrence depth but was removed from the soil following the careful removal of the wheat roots. Most of the roots of the plants in the rhizotrons exposed to PVC grew the entire length of the rhizotrons; however, fewer PVC particles were attached to the roots. These roots had an average surface area increase of 21% compared to the control. PVC was likely effectively incorporated into the soil matrix, as de Souza Machado et al. [41] demonstrated. Rillig et al. [21] suggest that the intensity of entanglement of microplastics in the planting medium would be higher in fibres, and fragments would experience greater mobility [22]; [23]; [24]. Jiang et al. [42] exposed the root tips of Broad bean Vicia faba to fluorescent polystyrene microplastics and discerned an attachment to the roots, although there was a decrease in their length.

Longer and finer roots were observed in the plants exposed to polyester. Polyester fibres have been evidenced to allow for good soil porosity, promoting root growth [41]. de Souza Machado et al. [41] observed that Allium fistulosum (spring onion) exposed to polyester fibres leads to lowered soil bulk density, thus potentially directly increasing root penetration ability with more viable space for the roots to grow within and improving soil aeration, which could lead to increased root growth [36]; [43]. As shown in Fig. 8, the average root biomass was highest in the roots exposed to polyester at 0.83 g, followed by 0.74 g for roots exposed to PVC and 0.56 g in the control group (Fig. S9). Root biomass increases more in soil with polyester fibres (48%) than in soilwith PVC fragments (32%) compared to he control group. This can be linked to reduced soil bulk density and increased soil microporosity. Increasing root mass would promote water and nutrient uptake, rhizodeposition, microbial activity, and mycorrhizal associations, which would ultimately improve aeration [40]; [36], and facilitate penetration of roots into the soil subsurface [43]. Thicker roots with low fineness would support quicker nutrient acquisition [44]; [45]. Root biomass was significantly increased with polyester fibres than PVC fragments, suggesting differential responses based on the shape of the microplastic [46]; [40].

Previous studies deduced conflicting results. Bosker et al. [47] observed an increase in root growth of Cress seeds (Lepidium Sativum) after exposure to an increasing size of microplastics due to adherence to the root hairs. In contrast, a reduction in root growth of aquatic duckweed species Spirodela polyrhiza [48] and Lemna minor [19] was observed when exposed to microplastics. Dovidat et al. [48] and Kalčíková et al. [19] deduced that the decreased growth could be attributed to the particles physically obstructing the roots. Comparable to this study, Lian et al. [49] ascertained that Triticum aestivum exposed to polystyrene nanoplastics promoted root elongation, thus increasing plant biomass with a reduced root-to-shoot biomass ratio. Increased root biomass in plants exposed to polyester increased the root-to-shoot ratio in the study by de Souza Machado et al. [40]. Consequently, the root-to-shoot ratio was highest in plants exposed to polyester at 5, 4 for plants exposed to PVC and 3 for plants in the control group. Therefore, it can be concluded that microplastic exposure increases the root-to-shoot ratio of wheat plants, with polyester having a noticeably higher increase. Furthermore, the root average diameter was quantifiably similar in roots in the control and those exposed to microplastics. Lehmann et al. [50] and de Souza Machado et al. [40] observed the contrary, where a decrease in the root average diameter in the presence of polyester fibres was found. A decrease in diameter in the presence of microplastics was hypothesized as a result of roots branching and overcoming root penetration resistance [32].

In conclusion, the impact of microplastics on wheat plants is multifaceted, affecting various aspects of plant growth and development. The physical effects include impedance of root growth resulting in increased root branching, possible root damage, and possible alterations in soil structure and aeration as demonstrated by Bao et al. [32], Bengough, A. G. [33], Rillig et al. [36]. , Boots et al. [38], and de Souza Machado et al. [41]. Chemical effects could encompass nutrient binding, additives leaching, and water flow disruption which is evident in Rillig et al. [36]. and de Souza Machado et al. [40]. Furthermore, microplastics can disrupt the soil microbiome and induce plant stress responses, leading to smaller leaves and oxidative stress as shown by Lozano et al. [46]. While polyester fibres likely promoted root growth by improving soil porosity and reducing bulk density, the higher water content in PVC-exposed plants might indicate an adaptive stress response. Like polyester fibres, other fibrous microplastics (e.g., polypropylene fibres) are likely to increase root surface area and promote fibrous root growth. Furthermore, hydrophobic microplastics like PVC tend to repel water and may not adhere firmly to root surfaces, while hydrophilic microplastics (e.g., nylon) might show different patterns of interaction and movement. Fragmented microplastics (e.g., polystyrene fragments) may not adhere as strongly to roots as fibres, leading to a moderate increase in root surface area and growth. Both polyester and PVC microplastics infiltrated deeper into the soil, especially with root growth exhibiting that roots facilitate the vertical movement of microplastics, potentially affecting deeper soil layers and groundwater.

Limitations of the experiment and future perspectives

This study explored how the shape of microplastics affects their movement into the soil through wheat root growth and how wheat responds to being exposed to microplastics. The experiment was conducted in controlled conditions, which might not fully represent real-world scenarios. The study focused on wheat (Triticum aestivum) and two types of microplastics (polyester fibres and PVC fragments). Still, it did not consider the potential interactions and effects that different crops and types of microplastics might have in various agricultural settings. The experiment utilized a concentration of microplastics (0.24% $\:w/w$ dry soil weight) at the upper limit of the range commonly found in natural agricultural fields, potentially exaggerating the observed effects. The study also primarily focused on the physical presence of microplastics on roots and did not extensively explore the biochemical interactions or long-term effects. Future research should include field studies under natural conditions to better understand the impacts of microplastic pollution on soil health and crop productivity. Additionally, investigating how microplastics change over time (e.g. ageing, fragmentation or biofilm formation) and their biochemical interactions with soil microorganisms and plant roots is crucial for understanding the broader ecological impacts of microplastic pollution.

Conclusion

Water infiltration and soil fauna have been documented to move microplastics along the soil profile vertically. This study investigated the role of plant root growth and elongation in the vertical movement of different shapes of microplastics in rhizotrons. Polyester fibres experienced infiltration into the soil due to strong adherence to roots; therefore, they move with root growth, elongation, and expansion. A direct relationship between root growth and microplastic fibre transport and infiltration can, therefore, be deduced. Moreover, phytoremediation can be evidenced, where during thinning of the seedlings after 7 days, their roots verifiably captured and effectively removed some polyester fibres from the sand-microplastic mix layer (0–20 cm) where they were planted. PVC fragments could have infiltrated through the fissures and pores created by root growth, elongation, preferential flow paths, and the water movement itself. Few PVC fragments adhered to the roots, and in the presence or absence of plant roots, a quantifiably similar number of fragments infiltrated up to 70 cm. Therefore, it can be concluded that vertical movement of differently shaped microplastics by the roots of the wheat plant (Triticum aestivum) in sandy soil is experienced as either of two scenarios. Firstly, strong adherence results in their movement with root growth and elongation, as seen in soil with polyester fibres. Secondly, through preferential flow paths in soil pores and the fissures created by root elongation experienced by PVC fragments. Control rhizotrons without wheat exhibited a more uniform distribution of microplastics in the upper 20 cm, implying that roots play a crucial role in influencing the vertical distribution of microplastics. While the shape of the microplastics (fibrous, e.g., polypropylene fibres and fragmented, e.g., polystyrene fragments and tire wear particles) governed their movement and interaction with the soil particles and roots, the plant type and its root growth pattern (extensive, fibrous root systems, e.g., grasses and cereals) had a significant impact on the overall behaviour of the microplastics in the soil. Together, these two factors determined the extent of microplastic infiltration and adherence in the soil profile, thereby shaping the experimental results. Additionally, hydrological conditions, biofilm formation, and microplastic ageing were not directly measured, but they may have an impact. Water infiltration is crucial for vertical infiltration. Biofilm formation and microplastic ageing change the surface properties, increasing adhesion to soil particles or roots, affecting infiltration.

This study was conducted under controlled conditions, not fully replicating natural field conditions. Future research should include field studies on the potential interactions and effects of different crops and types of microplastics at varied concentrations to validate these findings and understand the chronic effects of microplastic accumulation on soil health and crop productivity. This will provide a more comprehensive understanding of how microplastics impact soil and plant systems.